Nonradioactive alternative to clinical mixed lymphocyte reaction

Abstract

The MLR is used clinically as a functional assay for genetic HLA disparity. Traditionally this test relies on [ 3H]thymidine incorporation to detect T-cell proliferation as an indicator of alloantigenicity. Environmental and administrative concerns regarding radioisotope use and disposal have encouraged the development of sensitive, nonradioactive assay for T-cell alloactivation. We describe a nonradioactive alternative to the clinical MLR which uses an IL-2-dependent cell line, CTLL20.3, and MTT reduction to detect IL-2 accumulation in MLC SNs as an index of T-cell alloactivation. We first determined (a) the optimal number of CTLL20.3 cells for the assay, (b) the optimal time of SN analysis for IL-2, and (c) additional manipulations that significantly increase the assay sensitivity for IL-2. Using this assay system with patient lymphocyte combinations, we demonstrated that the CTLL20.3-MTT assays correlate well with the [ 3H]thymidine assays of T-cell proliferation for the detection of MHC incompatibility. Indeed, the CTLL20.3-MTT assay may be slightly more sensitive than the traditional clinical MLR. Human Immunology 43:38–44 (1995)